Development of reverse transcriptase polymerase chain reaction enzyme-linked immunosorbent assay for the detection of hepatitis A virus in vegetables

Abstract

This study's aim was to develop a specific and sensitive reverse transcription polymerase chain reaction enzyme-linked immunosorbent assay (RT-PCR-ELISA) for hepatitis A virus (HAV). Three concentration methods were compared using samples from cabbage, lettuce, and sesame leaves that were artificially contaminated with HAV. The specificity of the assay was tested against human norovirus, hepatitis E virus, rotavirus, enterovirus, and FCV. The sensitivity and specificity of RT-PCR-ELISA that were targeted to the 5'NCR and VP1 regions of HAV were compared. Nonspecific reactions were not observed. An optimal primer and probe set for RT-PCR-ELISA was selected for each VP1 and 5'NCR. The detection limit of the RT-PCR-ELISA was enhanced by 10-100 fold more than nested RT-PCR. Our new RT-PCR-ELISA was successfully optimized to screen vegetables for HAV contamination with high sensitivity in large samples. (C) 2011 Elsevier Ltd. All rights reserved.